2-oxo-1,2-dihydro-quinoline modulators of immune function

ABSTRACT

The present invention relates to new 2-oxo-1,2-dihydro-quinoline modulators of immune function, pharmaceutical compositions thereof, and methods of use thereof.

This application claims the benefit of priority of U.S. provisionalapplication No. 61/093,943, filed Sep. 3, 2008, the disclosure of whichis hereby incorporated by reference as if written herein in itsentirety.

Disclosed herein are new substituted 2-oxo-1,2-dihydro-quinolinecompounds, pharmaceutical compositions made thereof, and methods tomodulate immune function activity in a subject are also provided for,for the treatment of disorders such as multiple sclerosis and autoimmunedisorders.

Laquinimod (ABR 215062; SAIK-MS; ABR-215062; SAIKMS; CAS #248281-84-7),5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydro-quinoline-3-carboxylicacid ethyl-phenyl-amide, is an immune function modulator. Laquinimod iscurrently under investigation for the treatment of multiple sclerosis(Burton et al., Curr. Neurol. & Neurosc. Reports 2007, 7(3), 223-30;Tuvesson et al., Xenobiotica 2005, 35(3), 293-304; Cohen et al., Int. J.Clin. Pract. 2007, 61(11), 1922-30). Laquinimod has also shown promisein treating autoimmune disorders (Tuvesson et al., Xenobiotica 2005,35(3), 293-304).

Laquinimod is subject to extensive oxidative metabolism by cytochromeP₄₅₀ enzymes, particularly by CYP3A4 (Tuvesson et al., Drug Metab. &Disp. 2005, 33(6), 866-72). Primary metabolites include those formed byquinoline hydroxylation at various sites, quinoline demethylation,aniline de-ethylation, and aniline hydroxylation at the para position(Tuvesson et al., Xenobiotica 2005, 35(3), 293-304).

Deuterium Kinetic Isotope Effect

In order to eliminate foreign substances such as therapeutic agents, theanimal body expresses various enzymes, such as the cytochrome P₄₅₀enzymes (CYPs), esterases, proteases, reductases, dehydrogenases, andmonoamine oxidases, to react with and convert these foreign substancesto more polar intermediates or metabolites for renal excretion. Suchmetabolic reactions frequently involve the oxidation of acarbon-hydrogen (C—H) bond to either a carbon-oxygen (C—O) or acarbon-carbon (C—C) π-bond. The resultant metabolites may be stable orunstable under physiological conditions, and can have substantiallydifferent pharmacokinetic, pharmacodynamic, and acute and long-termtoxicity profiles relative to the parent compounds. For most drugs, suchoxidations are generally rapid and ultimately lead to administration ofmultiple or high daily doses.

The relationship between the activation energy and the rate of reactionmay be quantified by the Arrhenius equation, k=Ae^(−Eact/RT). TheArrhenius equation states that, at a given temperature, the rate of achemical reaction depends exponentially on the activation energy(E_(act)).

The transition state in a reaction is a short lived state along thereaction pathway during which the original bonds have stretched to theirlimit. By definition, the activation energy E_(act) for a reaction isthe energy required to reach the transition state of that reaction. Oncethe transition state is reached, the molecules can either revert to theoriginal reactants, or form new bonds giving rise to reaction products.A catalyst facilitates a reaction process by lowering the activationenergy leading to a transition state. Enzymes are examples of biologicalcatalysts.

Carbon-hydrogen bond strength is directly proportional to the absolutevalue of the ground-state vibrational energy of the bond. Thisvibrational energy depends on the mass of the atoms that form the bond,and increases as the mass of one or both of the atoms making the bondincreases. Since deuterium (D) has twice the mass of protium (¹H), a C-Dbond is stronger than the corresponding C—¹H bond. If a C—¹H bond isbroken during a rate-determining step in a chemical reaction (i.e. thestep with the highest transition state energy), then substituting adeuterium for that protium will cause a decrease in the reaction rate.This phenomenon is known as the Deuterium Kinetic Isotope Effect (DKIE).The magnitude of the DKIE can be expressed as the ratio between therates of a given reaction in which a C—¹H bond is broken, and the samereaction where deuterium is substituted for protium. The DKIE can rangefrom about 1 (no isotope effect) to very large numbers, such as 50 ormore. Substitution of tritium for hydrogen results in yet a strongerbond than deuterium and gives numerically larger isotope effects.

Deuterium (²H or D) is a stable and non-radioactive isotope of hydrogenwhich has approximately twice the mass of protium (¹H), the most commonisotope of hydrogen. Deuterium oxide (D₂O or “heavy water”) looks andtastes like H₂O, but has different physical properties.

When pure D₂O is given to rodents, it is readily absorbed. The quantityof deuterium required to induce toxicity is extremely high. When about0-15% of the body water has been replaced by D₂O, animals are healthybut are unable to gain weight as fast as the control (untreated) group.When about 15-20% of the body water has been replaced with D₂O, theanimals become excitable. When about 20-25% of the body water has beenreplaced with D₂O, the animals become so excitable that they go intofrequent convulsions when stimulated. Skin lesions, ulcers on the pawsand muzzles, and necrosis of the tails appear. The animals also becomevery aggressive. When about 30% of the body water has been replaced withD₂O, the animals refuse to eat and become comatose. Their body weightdrops sharply and their metabolic rates drop far below normal, withdeath occurring at about 30 to about 35% replacement with D₂O. Theeffects are reversible unless more than thirty percent of the previousbody weight has been lost due to D₂O. Studies have also shown that theuse of D₂O can delay the growth of cancer cells and enhance thecytotoxicity of certain antineoplastic agents.

Deuteration of pharmaceuticals to improve pharmacokinetics (PK),pharmacodynamics (PD), and toxicity profiles has been demonstratedpreviously with some classes of drugs. For example, the DKIE was used todecrease the hepatotoxicity of halothane, presumably by limiting theproduction of reactive species such as trifluoroacetyl chloride.However, this method may not be applicable to all drug classes. Forexample, deuterium incorporation can lead to metabolic switching.Metabolic switching occurs when xenogens, sequestered by Phase Ienzymes, bind transiently and re-bind in a variety of conformationsprior to the chemical reaction (e.g., oxidation). Metabolic switching isenabled by the relatively vast size of binding pockets in many Phase Ienzymes and the promiscuous nature of many metabolic reactions.Metabolic switching can lead to different proportions of knownmetabolites as well as altogether new metabolites. This new metabolicprofile may impart more or less toxicity. Such pitfalls are non-obviousand are not predictable a priori for any drug class.

Laquinimod is an immune function modulator. The carbon-hydrogen bonds oflaquinimod contain a naturally occurring distribution of hydrogenisotopes, namely ¹H or protium (about 99.9844%), ²H or deuterium (about0.0156%), and ³H or tritium (in the range between about 0.5 and 67tritium atoms per 10¹⁸ protium atoms). Increased levels of deuteriumincorporation may produce a detectable Deuterium Kinetic Isotope Effect(DKIE) that could effect the pharmacokinetic, pharmacologic and/ortoxicologic profiles of laquinimod in comparison with laquinimod havingnaturally occurring levels of deuterium.

Based on discoveries made in our laboratory, as well as considering theliterature, laquinimod is metabolized in humans at the quinoline ring,the N-methyl group, the N-ethyl group, and the phenyl ring. The currentapproach has the potential to prevent metabolism at these sites. Othersites on the molecule may also undergo transformations leading tometabolites with as-yet-unknown pharmacology/toxicology. Limiting theproduction of these metabolites has the potential to decrease the dangerof the administration of such drugs and may even allow increased dosageand/or increased efficacy. All of these transformations can occurthrough polymorphically-expressed enzymes, exacerbating interpatientvariability. Further, some disorders are best treated when the subjectis medicated around the clock or for an extended period of time. For allof the foregoing reasons, a medicine with a longer half-life may resultin greater efficacy and cost savings. Various deuteration patterns canbe used to (a) reduce or eliminate unwanted metabolites, (b) increasethe half-life of the parent drug, (c) decrease the number of dosesneeded to achieve a desired effect, (d) decrease the amount of a doseneeded to achieve a desired effect, (e) increase the formation of activemetabolites, if any are formed, (f) decrease the production ofdeleterious metabolites in specific tissues, and/or (g) create a moreeffective drug and/or a safer drug for polypharmacy, whether thepolypharmacy be intentional or not. The deuteration approach has thestrong potential to slow the metabolism of laquinimod and attenuateinterpatient variability.

Novel compounds and pharmaceutical compositions, certain of which havebeen found to modulate immune function have been discovered, togetherwith methods of synthesizing and using the compounds, including methodsfor the treatment of immune function-mediated disorders in a patient byadministering the compounds.

In certain embodiments of the present invention, compounds havestructural Formula I:

or a salt, solvate, or prodrug thereof, wherein:

R₁-R₁₇ are independently selected from the group consisting of hydrogenand deuterium; and

at least one of R₁-R₁₇ is deuterium.

Certain compounds disclosed herein may possess useful immune functionmodulating activity, and may be used in the treatment or prophylaxis ofa disorder in which immune function plays an active role. Thus, certainembodiments also provide pharmaceutical compositions comprising one ormore compounds disclosed herein together with a pharmaceuticallyacceptable carrier, as well as methods of making and using the compoundsand compositions. Certain embodiments provide methods for modulatingimmune function. Other embodiments provide methods for treating a immunefunction-mediated disorder in a patient in need of such treatment,comprising administering to said patient a therapeutically effectiveamount of a compound or composition according to the present invention.Also provided is the use of certain compounds disclosed herein for usein the manufacture of a medicament for the prevention or treatment of adisorder ameliorated by the modulation of immune function.

The compounds as disclosed herein may also contain less prevalentisotopes for other elements, including, but not limited to, ¹³C or ¹⁴Cfor carbon, ³³S, ³⁴S, or ³⁶S for sulfur, ¹⁵N for nitrogen, and ¹⁷O or¹⁸O for oxygen.

In certain embodiments, the compound disclosed herein may expose apatient to a maximum of about 0.000005% D₂O or about 0.00001% DHO,assuming that all of the C-D bonds in the compound as disclosed hereinare metabolized and released as D₂O or DHO. In certain embodiments, thelevels of D₂O shown to cause toxicity in animals is much greater thaneven the maximum limit of exposure caused by administration of thedeuterium enriched compound as disclosed herein. Thus, in certainembodiments, the deuterium-enriched compound disclosed herein should notcause any additional toxicity due to the formation of D₂O or DHO upondrug metabolism.

In certain embodiments, the deuterated compounds disclosed hereinmaintain the beneficial aspects of the corresponding non-isotopicallyenriched molecules while substantially increasing the maximum tolerateddose, decreasing toxicity, increasing the half-life (T_(1/2)), loweringthe maximum plasma concentration (C_(max)) of the minimum efficaciousdose (MED), lowering the efficacious dose and thus decreasing thenon-mechanism-related toxicity, and/or lowering the probability ofdrug-drug interactions.

All publications and references cited herein are expressly incorporatedherein by reference in their entirety. However, with respect to anysimilar or identical terms found in both the incorporated publicationsor references and those explicitly put forth or defined in thisdocument, then those terms definitions or meanings explicitly put forthin this document shall control in all respects.

As used herein, the terms below have the meanings indicated.

The singular forms “a,” “an,” and “the” may refer to plural articlesunless specifically stated otherwise.

The term “about,” as used herein, is intended to qualify the numericalvalues which it modifies, denoting such a value as variable within amargin of error. When no particular margin of error, such as a standarddeviation to a mean value given in a chart or table of data, is recited,the term “about” should be understood to mean that range which wouldencompass the recited value and the range which would be included byrounding up or down to that figure as well, taking into accountsignificant figures.

When ranges of values are disclosed, and the notation “from n₁ . . . ton₂” or “n₁-n₂” is used, where n₁ and n₂ are the numbers, then unlessotherwise specified, this notation is intended to include the numbersthemselves and the range between them. This range may be integral orcontinuous between and including the end values.

The term “deuterium enrichment” refers to the percentage ofincorporation of deuterium at a given position in a molecule in theplace of hydrogen. For example, deuterium enrichment of 1% at a givenposition means that 1% of molecules in a given sample contain deuteriumat the specified position. Because the naturally occurring distributionof deuterium is about 0.0156%, deuterium enrichment at any position in acompound synthesized using non-enriched starting materials is about0.0156%. The deuterium enrichment can be determined using conventionalanalytical methods known to one of ordinary skill in the art, includingmass spectrometry and nuclear magnetic resonance spectroscopy.

The term “is/are deuterium,” when used to describe a given position in amolecule such as R₁-R₁₇ or the symbol “D,” when used to represent agiven position in a drawing of a molecular structure, means that thespecified position is enriched with deuterium above the naturallyoccurring distribution of deuterium. In one embodiment deuteriumenrichment is no less than about 1%, in another no less than about 5%,in another no less than about 10%, in another no less than about 20%, inanother no less than about 50%, in another no less than about 70%, inanother no less than about 80%, in another no less than about 90%, or inanother no less than about 98% of deuterium at the specified position.

The term “isotopic enrichment” refers to the percentage of incorporationof a less prevalent isotope of an element at a given position in amolecule in the place of the more prevalent isotope of the element.

The term “non-isotopically enriched” refers to a molecule in which thepercentages of the various isotopes are substantially the same as thenaturally occurring percentages.

Asymmetric centers exist in the compounds disclosed herein. Thesecenters are designated by the symbols “R” or “S,” depending on theconfiguration of substituents around the chiral carbon atom. It shouldbe understood that the invention encompasses all stereochemical isomericforms, including diastereomeric, enantiomeric, and epimeric forms, aswell as D-isomers and L-isomers, and mixtures thereof. Individualstereoisomers of compounds can be prepared synthetically fromcommercially available starting materials which contain chiral centersor by preparation of mixtures of enantiomeric products followed byseparation such as conversion to a mixture of diastereomers followed byseparation or recrystallization, chromatographic techniques, directseparation of enantiomers on chiral chromatographic columns, or anyother appropriate method known in the art. Starting compounds ofparticular stereochemistry are either commercially available or can bemade and resolved by techniques known in the art. Additionally, thecompounds disclosed herein may exist as geometric isomers. The presentinvention includes all cis, trans, syn, anti, entgegen (E), and zusammen(Z) isomers as well as the appropriate mixtures thereof. Additionally,compounds may exist as tautomers; all tautomeric isomers are provided bythis invention. Additionally, the compounds disclosed herein can existin unsolvated as well as solvated forms with pharmaceutically acceptablesolvents such as water, ethanol, and the like. In general, the solvatedforms are considered equivalent to the unsolvated forms.

The term “bond” refers to a covalent linkage between two atoms, or twomoieties when the atoms joined by the bond are considered to be part oflarger substructure. A bond may be single, double, or triple unlessotherwise specified. A dashed line between two atoms in a drawing of amolecule indicates that an additional bond may be present or absent atthat position.

The term “disorder” as used herein is intended to be generallysynonymous, and is used interchangeably with, the terms “disease” and“condition” (as in medical condition), in that all reflect an abnormalcondition of the human or animal body or of one of its parts thatimpairs normal functioning, is typically manifested by distinguishingsigns and symptoms.

The terms “treat,” “treating,” and “treatment” are meant to includealleviating or abrogating a disorder or one or more of the symptomsassociated with a disorder; or alleviating or eradicating the cause(s)of the disorder itself. As used herein, reference to “treatment” of adisorder is intended to include prevention. The terms “prevent,”“preventing,” and “prevention” refer to a method of delaying orprecluding the onset of a disorder; and/or its attendant symptoms,barring a subject from acquiring a disorder or reducing a subject's riskof acquiring a disorder.

The term “therapeutically effective amount” refers to the amount of acompound that, when administered, is sufficient to prevent developmentof, or alleviate to some extent, one or more of the symptoms of thedisorder being treated. The term “therapeutically effective amount” alsorefers to the amount of a compound that is sufficient to elicit thebiological or medical response of a cell, tissue, system, animal, orhuman that is being sought by a researcher, veterinarian, medicaldoctor, or clinician.

The term “subject” refers to an animal, including, but not limited to, aprimate (e.g., human, monkey, chimpanzee, gorilla, and the like),rodents (e.g., rats, mice, gerbils, hamsters, ferrets, and the like),lagomorphs, swine (e.g., pig, miniature pig), equine, canine, feline,and the like. The terms “subject” and “patient” are used interchangeablyherein in reference, for example, to a mammalian subject, such as ahuman patient.

The term “combination therapy” means the administration of two or moretherapeutic agents to treat a therapeutic disorder described in thepresent disclosure. Such administration encompasses co-administration ofthese therapeutic agents in a substantially simultaneous manner, such asin a single capsule having a fixed ratio of active ingredients or inmultiple, separate capsules for each active ingredient. In addition,such administration also encompasses use of each type of therapeuticagent in a sequential manner. In either case, the treatment regimen willprovide beneficial effects of the drug combination in treating thedisorders described herein.

The term “immune function” refers to the collection of mechanisms withinan organism that protects against disease. Such mechanisms includemacrophages, T-lymphocytes, and B-lymphocytes and their respectiveactivities.

The term “immune function-mediated disorder,” refers to a disorder thatis characterized by abnormal immune function. An immunefunction-mediated disorder may be completely or partially mediated bymodulating the immune function in a subject. In particular, an immunefunction-mediated disorder is one in which modulation of immune functionresults in some effect on the underlying disorder e.g., administrationof a immune function modulator results in some improvement in at leastsome of the patients being treated.

The term “immune function modulator,” refers to the ability of acompound disclosed herein to alter immune function activity. An immunefunction modulator may stimulate immune function activity, may activateor inhibit immune function activity depending on the concentration ofthe compound exposed to the subject, or may inhibit immune functionactivity. Such activation or inhibition may be contingent on theoccurrence of a specific event, such as activation of a signaltransduction pathway, and/or may be manifest only in particular celltypes. For example, compounds disclosed herein may modulate immunefunction by inhibiting the infiltration of both CD4⁺ T-cells andmacrophages into central nervous tissues and changing the T-lymphocytepopulation in favour of cells expressing Th2/Th3 cytokines interleukin(IL)-4, IL-10 and transforming growth factor-beta. In some embodiments,modulation of the immune function may be assessed using the methoddescribed in Karussis et al., Ann. Neurol. 1993, (34), 654-660; Yang, etal., Journal of Neuroimmunology 2004, 156(1-2), 3-9; Brunmark et al., J.Neuroimmunol. 2002, 130, 163-172; and Jonsson et al., J. Med. Chem.2004, 47, 2075-88.

The term “therapeutically acceptable” refers to those compounds (orsalts, prodrugs, tautomers, zwitterionic forms, etc.) which are suitablefor use in contact with the tissues of patients without excessivetoxicity, irritation, allergic response, immunogenecity, arecommensurate with a reasonable benefit/risk ratio, and are effective fortheir intended use.

The term “pharmaceutically acceptable carrier,” “pharmaceuticallyacceptable excipient,” “physiologically acceptable carrier,” or“physiologically acceptable excipient” refers to apharmaceutically-acceptable material, composition, or vehicle, such as aliquid or solid filler, diluent, excipient, solvent, or encapsulatingmaterial. Each component must be “pharmaceutically acceptable” in thesense of being compatible with the other ingredients of a pharmaceuticalformulation. It must also be suitable for use in contact with the tissueor organ of humans and animals without excessive toxicity, irritation,allergic response, immunogenecity, or other problems or complications,commensurate with a reasonable benefit/risk ratio. See, Remington: TheScience and Practice of Pharmacy, 21st Edition; Lippincott Williams &Wilkins: Philadelphia, Pa., 2005; Handbook of Pharmaceutical Excipients,5th Edition; Rowe et al., Eds., The Pharmaceutical Press and theAmerican Pharmaceutical Association: 2005; and Handbook ofPharmaceutical Additives, 3rd Edition; Ash and Ash Eds., GowerPublishing Company: 2007; Pharmaceutical Preformulation and Formulation,Gibson Ed., CRC Press LLC: Boca Raton, Fla., 2004).

The terms “active ingredient,” “active compound,” and “active substance”refer to a compound, which is administered, alone or in combination withone or more pharmaceutically acceptable excipients or carriers, to asubject for treating, preventing, or ameliorating one or more symptomsof a disorder.

The terms “drug,” “therapeutic agent,” and “chemotherapeutic agent”refer to a compound, or a pharmaceutical composition thereof, which isadministered to a subject for treating, preventing, or ameliorating oneor more symptoms of a disorder.

The term “release controlling excipient” refers to an excipient whoseprimary function is to modify the duration or place of release of theactive substance from a dosage form as compared with a conventionalimmediate release dosage form.

The term “nonrelease controlling excipient” refers to an excipient whoseprimary function do not include modifying the duration or place ofrelease of the active substance from a dosage form as compared with aconventional immediate release dosage form.

The term “prodrug” refers to a compound functional derivative of thecompound as disclosed herein and is readily convertible into the parentcompound in vivo. Prodrugs are often useful because, in some situations,they may be easier to administer than the parent compound. They may, forinstance, be bioavailable by oral administration whereas the parentcompound is not. The prodrug may also have enhanced solubility inpharmaceutical compositions over the parent compound. A prodrug may beconverted into the parent drug by various mechanisms, includingenzymatic processes and metabolic hydrolysis. See Harper, Progress inDrug Research 1962, 4, 221-294; Morozowich et al. in “Design ofBiopharmaceutical Properties through Prodrugs and Analogs,” Roche Ed.,APHA Acad. Pharm. Sci. 1977; “Bioreversible Carriers in Drug in DrugDesign, Theory and Application,” Roche Ed., APHA Acad. Pharm. Sci. 1987;“Design of Prodrugs,” Bundgaard, Elsevier, 1985; Wang et al., Curr.Pharm. Design 1999, 5, 265-287; Pauletti et al., Adv. Drug. DeliveryRev. 1997, 27, 235-256; Mizen et al., Pharm. Biotech. 1998, 11, 345-365;Gaignault et al., Pract. Med. Chem. 1996, 671-696; Asgharnejad in“Transport Processes in Pharmaceutical Systems,” Amidon et al., Ed.,Marcell Dekker, 185-218, 2000; Balant et al., Eur. J. Drug Metab.Pharmacokinet. 1990, 15, 143-53; Balimane and Sinko, Adv. Drug DeliveryRev. 1999, 39, 183-209; Browne, Clin. Neuropharmacol. 1997, 20, 1-12;Bundgaard, Arch. Pharm. Chem. 1979, 86, 1-39; Bundgaard, Controlled DrugDelivery 1987, 17, 179-96; Bundgaard, Adv. Drug Delivery Rev. 1992, 8,1-38; Fleisher et al., Adv. Drug Delivery Rev. 1996, 19, 115-130;Fleisher et al., Methods Enzymol. 1985, 112, 360-381; Farquhar et al.,J. Pharm. Sci. 1983, 72, 324-325; Freeman et al., J. Chem. Soc., Chem.Commun. 1991, 875-877; Friis and Bundgaard, Eur. J. Pharm. Sci. 1996, 4,49-59; Gangwar et al., Des. Biopharm. Prop. Prodrugs Analogs, 1977,409-421; Nathwani and Wood, Drugs 1993, 45, 866-94; Sinhababu andThakker, Adv. Drug Delivery Rev. 1996, 19, 241-273; Stella et al., Drugs1985, 29, 455-73; Tan et al., Adv. Drug Delivery Rev. 1999, 39, 117-151;Taylor, Adv. Drug Delivery Rev. 1996, 19, 131-148; Valentino andBorchardt, Drug Discovery Today 1997, 2, 148-155; Wiebe and Knaus, Adv.Drug Delivery Rev. 1999, 39, 63-80; Waller et al., Br. J. Clin. Pharmac.1989, 28, 497-507.

The compounds disclosed herein can exist as therapeutically acceptablesalts. The term “therapeutically acceptable salt,” as used herein,represents salts or zwitterionic forms of the compounds disclosed hereinwhich are therapeutically acceptable as defined herein. The salts can beprepared during the final isolation and purification of the compounds orseparately by reacting the appropriate compound with a suitable acid orbase. Therapeutically acceptable salts include acid and basic additionsalts. For a more complete discussion of the preparation and selectionof salts, refer to “Handbook of Pharmaceutical Salts, Properties, andUse,” Stah and Wermuth, Ed.; (Wiley-VCH and VHCA, Zurich, 2002) andBerge et al., J. Pharm. Sci. 1977, 66, 1-19. Suitable acids for use inthe preparation of pharmaceutically acceptable salts include, but arenot limited to, acetic acid, 2,2-dichloroacetic acid, acylated aminoacids, adipic acid, alginic acid, ascorbic acid, L-aspartic acid,benzenesulfonic acid, benzoic acid, 4-acetamidobenzoic acid, boric acid,(+)-camphoric acid, camphorsulfonic acid, (+)-(1S)-camphor-10-sulfonicacid, capric acid, caproic acid, caprylic acid, cinnamic acid, citricacid, cyclamic acid, cyclohexanesulfamic acid, dodecylsulfuric acid,ethane-1,2-disulfonic acid, ethanesulfonic acid,2-hydroxy-ethanesulfonic acid, formic acid, fumaric acid, galactaricacid, gentisic acid, glucoheptonic acid, D-gluconic acid, D-glucuronicacid, L-glutamic acid, α-oxo-glutaric acid, glycolic acid, hippuricacid, hydrobromic acid, hydrochloric acid, hydroiodic acid, (+)-L-lacticacid, (±)-DL-lactic acid, lactobionic acid, lauric acid, maleic acid,(−)-L-malic acid, malonic acid, (±)-DL-mandelic acid, methanesulfonicacid, naphthalene-2-sulfonic acid, naphthalene-1,5-disulfonic acid,1-hydroxy-2-naphthoic acid, nicotinic acid, nitric acid, oleic acid,orotic acid, oxalic acid, palmitic acid, pamoic acid, perchloric acid,phosphoric acid, L-pyroglutamic acid, saccharic acid, salicylic acid,4-amino-salicylic acid, sebacic acid, stearic acid, succinic acid,sulfuric acid, tannic acid, (+)-L-tartaric acid, thiocyanic acid,p-toluenesulfonic acid, undecylenic acid, and valeric acid.

Suitable bases for use in the preparation of pharmaceutically acceptablesalts, including, but not limited to, inorganic bases, such as magnesiumhydroxide, calcium hydroxide, potassium hydroxide, zinc hydroxide, orsodium hydroxide; and organic bases, such as primary, secondary,tertiary, and quaternary, aliphatic and aromatic amines, includingL-arginine, benethamine, benzathine, choline, deanol, diethanolamine,diethylamine, dimethylamine, dipropylamine, diisopropylamine,2-(diethylamino)-ethanol, ethanolamine, ethylamine, ethylenediamine,isopropylamine, N-methyl-glucamine, hydrabamine, 1H-imidazole, L-lysine,morpholine, 4-(2-hydroxyethyl)-morpholine, methylamine, piperidine,piperazine, propylamine, pyrrolidine, 1-(2-hydroxyethyl)-pyrrolidine,pyridine, quinuclidine, quinoline, isoquinoline, secondary amines,triethanolamine, trimethylamine, triethylamine, N-methyl-D-glucamine,2-amino-2-(hydroxymethyl)-1,3-propanediol, and tromethamine.

While it may be possible for the compounds of the subject invention tobe administered as the raw chemical, it is also possible to present themas a pharmaceutical composition. Accordingly, provided herein arepharmaceutical compositions which comprise one or more of certaincompounds disclosed herein, or one or more pharmaceutically acceptablesalts, prodrugs, or solvates thereof, together with one or morepharmaceutically acceptable carriers thereof and optionally one or moreother therapeutic ingredients. Proper formulation is dependent upon theroute of administration chosen. Any of the well-known techniques,carriers, and excipients may be used as suitable and as understood inthe art; e.g., in Remington's Pharmaceutical Sciences. Thepharmaceutical compositions disclosed herein may be manufactured in anymanner known in the art, e.g., by means of conventional mixing,dissolving, granulating, dragee-making, levigating, emulsifying,encapsulating, entrapping or compression processes. The pharmaceuticalcompositions may also be formulated as a modified release dosage form,including delayed-, extended-, prolonged-, sustained-, pulsatile-,controlled-, accelerated- and fast-, targeted-, programmed-release, andgastric retention dosage forms. These dosage forms can be preparedaccording to conventional methods and techniques known to those skilledin the art (see, Remington: The Science and Practice of Pharmacy, supra;Modified-Release Drug Deliver Technology, Rathbone et al., Eds., Drugsand the Pharmaceutical Science, Marcel Dekker, Inc.: New York, N.Y.,2002; Vol. 126).

The compositions include those suitable for oral, parenteral (includingsubcutaneous, intradermal, intramuscular, intravenous, intraarticular,and intramedullary), intraperitoneal, transmucosal, transdermal, rectaland topical (including dermal, buccal, sublingual and intraocular)administration although the most suitable route may depend upon forexample the condition and disorder of the recipient. The compositionsmay conveniently be presented in unit dosage form and may be prepared byany of the methods well known in the art of pharmacy. Typically, thesemethods include the step of bringing into association a compound of thesubject invention or a pharmaceutically salt, prodrug, or solvatethereof (“active ingredient”) with the carrier which constitutes one ormore accessory ingredients. In general, the compositions are prepared byuniformly and intimately bringing into association the active ingredientwith liquid carriers or finely divided solid carriers or both and then,if necessary, shaping the product into the desired formulation.

Formulations of the compounds disclosed herein suitable for oraladministration may be presented as discrete units such as capsules,cachets or tablets each containing a predetermined amount of the activeingredient; as a powder or granules; as a solution or a suspension in anaqueous liquid or a non-aqueous liquid; or as an oil-in-water liquidemulsion or a water-in-oil liquid emulsion. The active ingredient mayalso be presented as a bolus, electuary or paste.

Pharmaceutical preparations which can be used orally include tablets,push-fit capsules made of gelatin, as well as soft, sealed capsules madeof gelatin and a plasticizer, such as glycerol or sorbitol. Tablets maybe made by compression or molding, optionally with one or more accessoryingredients. Compressed tablets may be prepared by compressing in asuitable machine the active ingredient in a free-flowing form such as apowder or granules, optionally mixed with binders, inert diluents, orlubricating, surface active or dispersing agents. Molded tablets may bemade by molding in a suitable machine a mixture of the powdered compoundmoistened with an inert liquid diluent. The tablets may optionally becoated or scored and may be formulated so as to provide slow orcontrolled release of the active ingredient therein. All formulationsfor oral administration should be in dosages suitable for suchadministration. The push-fit capsules can contain the active ingredientsin admixture with filler such as lactose, binders such as starches,and/or lubricants such as talc or magnesium stearate and, optionally,stabilizers. In soft capsules, the active compounds may be dissolved orsuspended in suitable liquids, such as fatty oils, liquid paraffin, orliquid polyethylene glycols. In addition, stabilizers may be added.Dragee cores are provided with suitable coatings. For this purpose,concentrated sugar solutions may be used, which may optionally containgum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethyleneglycol, and/or titanium dioxide, lacquer solutions, and suitable organicsolvents or solvent mixtures. Dyestuffs or pigments may be added to thetablets or dragee coatings for identification or to characterizedifferent combinations of active compound doses.

The compounds may be formulated for parenteral administration byinjection, e.g., by bolus injection or continuous infusion. Formulationsfor injection may be presented in unit dosage form, e.g., in ampoules orin multi-dose containers, with an added preservative. The compositionsmay take such forms as suspensions, solutions or emulsions in oily oraqueous vehicles, and may contain formulatory agents such as suspending,stabilizing and/or dispersing agents. The formulations may be presentedin unit-dose or multi-dose containers, for example sealed ampoules andvials, and may be stored in powder form or in a freeze-dried(lyophilized) condition requiring only the addition of the sterileliquid carrier, for example, saline or sterile pyrogen-free water,immediately prior to use. Extemporaneous injection solutions andsuspensions may be prepared from sterile powders, granules and tabletsof the kind previously described.

Formulations for parenteral administration include aqueous andnon-aqueous (oily) sterile injection solutions of the active compoundswhich may contain antioxidants, buffers, bacteriostats and solutes whichrender the formulation isotonic with the blood of the intendedrecipient; and aqueous and non-aqueous sterile suspensions which mayinclude suspending agents and thickening agents. Suitable lipophilicsolvents or vehicles include fatty oils such as sesame oil, or syntheticfatty acid esters, such as ethyl oleate or triglycerides, or liposomes.Aqueous injection suspensions may contain substances which increase theviscosity of the suspension, such as sodium carboxymethyl cellulose,sorbitol, or dextran. Optionally, the suspension may also containsuitable stabilizers or agents which increase the solubility of thecompounds to allow for the preparation of highly concentrated solutions.

In addition to the formulations described previously, the compounds mayalso be formulated as a depot preparation. Such long acting formulationsmay be administered by implantation (for example subcutaneously orintramuscularly) or by intramuscular injection. Thus, for example, thecompounds may be formulated with suitable polymeric or hydrophobicmaterials (for example as an emulsion in an acceptable oil) or ionexchange resins, or as sparingly soluble derivatives, for example, as asparingly soluble salt.

For buccal or sublingual administration, the compositions may take theform of tablets, lozenges, pastilles, or gels formulated in conventionalmanner. Such compositions may comprise the active ingredient in aflavored basis such as sucrose and acacia or tragacanth.

The compounds may also be formulated in rectal compositions such assuppositories or retention enemas, e.g., containing conventionalsuppository bases such as cocoa butter, polyethylene glycol, or otherglycerides.

Certain compounds disclosed herein may be administered topically, thatis by non-systemic administration. This includes the application of acompound disclosed herein externally to the epidermis or the buccalcavity and the instillation of such a compound into the ear, eye andnose, such that the compound does not significantly enter the bloodstream. In contrast, systemic administration refers to oral,intravenous, intraperitoneal and intramuscular administration.

Formulations suitable for topical administration include liquid orsemi-liquid preparations suitable for penetration through the skin tothe site of inflammation such as gels, liniments, lotions, creams,ointments or pastes, and drops suitable for administration to the eye,ear or nose.

For administration by inhalation, compounds may be delivered from aninsufflator, nebulizer pressurized packs or other convenient means ofdelivering an aerosol spray. Pressurized packs may comprise a suitablepropellant such as dichlorodifluoromethane, trichlorofluoromethane,dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In thecase of a pressurized aerosol, the dosage unit may be determined byproviding a valve to deliver a metered amount. Alternatively, foradministration by inhalation or insufflation, the compounds according tothe invention may take the form of a dry powder composition, for examplea powder mix of the compound and a suitable powder base such as lactoseor starch. The powder composition may be presented in unit dosage form,in for example, capsules, cartridges, gelatin or blister packs fromwhich the powder may be administered with the aid of an inhalator orinsufflator.

Preferred unit dosage formulations are those containing an effectivedose, as herein below recited, or an appropriate fraction thereof, ofthe active ingredient.

Compounds may be administered orally or via injection at a dose of from0.1 to 500 mg/kg per day. The dose range for adult humans is generallyfrom 5 mg to 2 g/day. Tablets or other forms of presentation provided indiscrete units may conveniently contain an amount of one or morecompounds which is effective at such dosage or as a multiple of thesame, for instance, units containing 5 mg to 500 mg, usually around 10mg to 200 mg.

The amount of active ingredient that may be combined with the carriermaterials to produce a single dosage form will vary depending upon thehost treated and the particular mode of administration.

The compounds can be administered in various modes, e.g. orally,topically, or by injection. The precise amount of compound administeredto a patient will be the responsibility of the attendant physician. Thespecific dose level for any particular patient will depend upon avariety of factors including the activity of the specific compoundemployed, the age, body weight, general health, sex, diets, time ofadministration, route of administration, rate of excretion, drugcombination, the precise disorder being treated, and the severity of thedisorder being treated. Also, the route of administration may varydepending on the disorder and its severity.

In the case wherein the patient's condition does not improve, upon thedoctor's discretion the administration of the compounds may beadministered chronically, that is, for an extended period of time,including throughout the duration of the patient's life in order toameliorate or otherwise control or limit the symptoms of the patient'sdisorder.

In the case wherein the patient's status does improve, upon the doctor'sdiscretion the administration of the compounds may be given continuouslyor temporarily suspended for a certain length of time (i.e., a “drugholiday”).

Once improvement of the patient's conditions has occurred, a maintenancedose is administered if necessary. Subsequently, the dosage or thefrequency of administration, or both, can be reduced, as a function ofthe symptoms, to a level at which the improved disorder is retained.Patients can, however, require intermittent treatment on a long-termbasis upon any recurrence of symptoms.

Disclosed herein are methods of treating an immune function-mediateddisorder comprising administering to a subject having or suspected tohave such a disorder, a therapeutically effective amount of a compoundas disclosed herein or a pharmaceutically acceptable salt, solvate, orprodrug thereof.

Immune function-mediated disorders, include, but are not limited to,multiple sclerosis and autoimmune disorders, and/or any disorder whichcan lessened, alleviated, or prevented by administering a immunefunction modulator.

In certain embodiments, a method of treating a immune function-mediateddisorder comprises administering to the subject a therapeuticallyeffective amount of a compound of as disclosed herein, or apharmaceutically acceptable salt, solvate, or prodrug thereof, so as toaffect: (1) decreased inter-individual variation in plasma levels of thecompound or a metabolite thereof, (2) increased average plasma levels ofthe compound or decreased average plasma levels of at least onemetabolite of the compound per dosage unit; (3) decreased inhibition of,and/or metabolism by at least one cytochrome P₄₅₀ or monoamine oxidaseisoform in the subject; (4) decreased metabolism via at least onepolymorphically-expressed cytochrome P₄₅₀ isoform in the subject; (5) atleast one statistically-significantly improved disorder-control and/ordisorder-eradication endpoint; (6) an improved clinical effect duringthe treatment of the disorder, (7) prevention of recurrence, or delay ofdecline or appearance, of abnormal alimentary or hepatic parameters asthe primary clinical benefit, or (8) reduction or elimination ofdeleterious changes in any diagnostic hepatobiliary function endpoints,as compared to the corresponding non-isotopically enriched compound.

In certain embodiments, inter-individual variation in plasma levels ofthe compounds as disclosed herein, or metabolites thereof, is decreased;average plasma levels of the compound as disclosed herein are increased;average plasma levels of a metabolite of the compound as disclosedherein are decreased; inhibition of a cytochrome P₄₅₀ or monoamineoxidase isoform by a compound as disclosed herein is decreased; ormetabolism of the compound as disclosed herein by at least onepolymorphically-expressed cytochrome P₄₅₀ isoform is decreased; bygreater than about 5%, greater than about 10%, greater than about 20%,greater than about 30%, greater than about 40%, or by greater than about50% as compared to the corresponding non-isotopically enriched compound.

Plasma levels of the compound as disclosed herein, or metabolitesthereof, may be measured using the methods described by Li et al. RapidCommunications in Mass Spectrometry 2005, 19, 1943-1950; Sennbro, etal., Rapid Communications in Mass Spectrometry 2006, 20(22), 3313-3318;Edman, et al., Journal of Chromatography, B: Analytical Technologies inthe Biomedical and Life Sciences 2003, 785(2); and any references citedtherein and modifications made thereof.

Examples of cytochrome P₄₅₀ isoforms in a mammalian subject include, butare not limited to, CYP1A1, CYP1A2, CYP1B1, CYP2A6, CYP2A13, CYP2B6,CYP2C8, CYP2C9, CYP2C18, CYP2C19, CYP2D6, CYP2E1, CYP2G1, CYP2J2,CYP2R1, CYP2S1, CYP3A4, CYP3A5, CYP3A5P1, CYP3A5P2, CYP3A7, CYP4A11,CYP4B1, CYP4F2, CYP4F3, CYP4F8, CYP4F11, CYP4F12, CYP4X1, CYP4Z1,CYP5A1, CYP7A1, CYP7B1, CYP8A1, CYP8B1, CYP11A1, CYP11B1, CYP11B2,CYP17, CYP19, CYP21, CYP24, CYP26A1, CYP26B1, CYP27A1, CYP27B1, CYP39,CYP46, and CYP51.

Examples of monoamine oxidase isoforms in a mammalian subject include,but are not limited to, MAO_(A), and MAO_(B).

The inhibition of the cytochrome P₄₅₀ isoform is measured by the methodof Ko et al., British Journal of Clinical Pharmacology, 2000, 49,343-351. The inhibition of the MAO_(A) isoform is measured by the methodof Weyler et al., J. Biol. Chem. 1985, 260, 13199-13207. The inhibitionof the MAO_(B) isoform is measured by the method of Uebelhack et al.Pharmacopsychiatry, 1998, 31, 187-192.

Examples of polymorphically-expressed cytochrome P₄₅₀ isoforms in amammalian subject include, but are not limited to, CYP2C8, CYP2C9,CYP2C19, and CYP2D6.

The metabolic activities of liver microsomes, cytochrome P₄₅₀ isoforms,and monoamine oxidase isoforms are measured by the methods describedherein.

Examples of improved disorder-control and/or disorder-eradicationendpoints, or improved clinical effects include, but are not limited to,cumulative number of active lesions seen at week 24, cumulative andactive number of active and gadolinium-enhancing lesions on MRI every 8weeks, relapse rate, multiple sclerosis functional composite, short form36 quality of life assessment (Burton et al., Curr. Neurol. & Neurosc.Reports 2007, 7(3), 223-30).

Examples of diagnostic hepatobiliary function endpoints include, but arenot limited to, alanine aminotransferase (“ALT”), serum glutamic-pyruvictransaminase (“SGPT”), aspartate aminotransferase (“AST” or “SGOT”),ALT/AST ratios, serum aldolase, alkaline phosphatase (“ALP”), ammonialevels, bilirubin, gamma-glutamyl transpeptidase (“GGTP,” “γ-GTP,” or“GGT”), leucine aminopeptidase (“LAP”), liver biopsy, liverultrasonography, liver nuclear scan, 5′-nucleotidase, and blood protein.Hepatobiliary endpoints are compared to the stated normal levels asgiven in “Diagnostic and Laboratory Test Reference”, 4^(th) edition,Mosby, 1999. These assays are run by accredited laboratories accordingto standard protocol.

Besides being useful for human treatment, certain compounds andformulations disclosed herein may also be useful for veterinarytreatment of companion animals, exotic animals and farm animals,including mammals, rodents, and the like. More preferred animals includehorses, dogs, and cats.

Combination Therapy

The compounds disclosed herein may also be combined or used incombination with other agents useful in the treatment of immunefunction-mediated disorders. Or, by way of example only, the therapeuticeffectiveness of one of the compounds described herein may be enhancedby administration of an adjuvant (i.e., by itself the adjuvant may onlyhave minimal therapeutic benefit, but in combination with anothertherapeutic agent, the overall therapeutic benefit to the patient isenhanced).

Such other agents, adjuvants, or drugs, may be administered, by a routeand in an amount commonly used therefor, simultaneously or sequentiallywith a compound as disclosed herein. When a compound as disclosed hereinis used contemporaneously with one or more other drugs, a pharmaceuticalcomposition containing such other drugs in addition to the compounddisclosed herein may be utilized, but is not required.

In certain embodiments, the compounds disclosed herein can be combinedwith one or more immunomodulators, steroidal drugs or cyclosporins.

In certain embodiments, the compounds provided herein can be combinedwith one or more immunomodulators known in the art, including, but notlimited to, filgrastim, molgramostim, sargramostim, lenograstim,ancestim, pegfilgrastim, interferon gamma, interferon alpha-2a,interferon alpha-2b, interferon alpha-n1, interferon beta-1a, interferonbeta-1b, interferon alphacon-1, peginterferon alpha-2b, peginterferonalpha-2a, interferon omega, aldesleukin, oprelvekin, lentinan,roquinimex, BCG vaccine, pegademase, pidotimod, Poly I:C, Poly ICLC,thymopentin, immunocyanin, tasonermin, melanoma vaccine, glatirameracetate, histamine dihydrochloride, mifamurtide, plerixafor,muromonab-CD3, antilymphocyte immunoglobulin (horse), antithymocyteimmunoglobulin (rabbit), mycophenolic acid, sirolimus, leflunomide,alefacept, everolimus, gusperimus, efalizumab, abetimus, natalizumab,abatacept, eculizumab, etanercept, infliximab, afelimomab, adalimumab,certolizumab pegol, daclizumab, basiliximab, anakinra, ciclosporin,tacrolimus, azathioprine, thalidomide, methotrexate, and lenalidomide.

The compounds disclosed herein can also be administered in combinationwith other classes of compounds, including, but not limited to,norepinephrine reuptake inhibitors (NRIs) such as atomoxetine; dopaminereuptake inhibitors (DARIs), such as methylphenidate;serotonin-norepinephrine reuptake inhibitors (SNRIs), such asmilnacipran; sedatives, such as diazepham; norepinephrine-dopaminereuptake inhibitor (NDRIs), such as bupropion;serotonin-norepinephrine-dopamine-reuptake-inhibitors (SNDRIs), such asvenlafaxine; monoamine oxidase inhibitors, such as selegiline;hypothalamic phospholipids; endothelin converting enzyme (ECE)inhibitors, such as phosphoramidon; opioids, such as tramadol;thromboxane receptor antagonists, such as ifetroban; potassium channelopeners; thrombin inhibitors, such as hirudin; hypothalamicphospholipids; growth factor inhibitors, such as modulators of PDGFactivity; platelet activating factor (PAF) antagonists; anti-plateletagents, such as GPIIb/IIIa blockers (e.g., abdximab, eptifibatide, andtirofiban), P2Y(AC) antagonists (e.g., clopidogrel, ticlopidine andCS-747), and aspirin; anticoagulants, such as warfarin; low molecularweight heparins, such as enoxaparin; Factor VIIa Inhibitors and FactorXa Inhibitors; renin inhibitors; neutral endopeptidase (NEP) inhibitors;vasopepsidase inhibitors (dual NEP-ACE inhibitors), such as omapatrilatand gemopatrilat; HMG CoA reductase inhibitors, such as pravastatin,lovastatin, atorvastatin, simvastatin, NK-104 (a.k.a. itavastatin,nisvastatin, or nisbastatin), and ZD-4522 (also known as rosuvastatin,or atavastatin or visastatin); squalene synthetase inhibitors; fibrates;bile acid sequestrants, such as questran; niacin; anti-atheroscleroticagents, such as ACAT inhibitors; MTP Inhibitors; calcium channelblockers, such as amlodipine besylate; potassium channel activators;alpha-muscarinic agents; beta-muscarinic agents, such as carvedilol andmetoprolol; antiarrhythmic agents; diuretics, such as chlorothlazide,hydrochlorothiazide, flumethiazide, hydroflumethiazide,bendroflumethiazide, methylchlorothiazide, trichloromethiazide,polythiazide, benzothlazide, ethacrynic acid, tricrynafen,chlorthalidone, furosenilde, musolimine, bumetanide, triamterene,amiloride, and spironolactone; thrombolytic agents, such as tissueplasminogen activator (tPA), recombinant tPA, streptokinase, urokinase,prourokinase, and anisoylated plasminogen streptokinase activatorcomplex (APSAC); anti-diabetic agents, such as biguanides (e.g.metformin), glucosidase inhibitors (e.g., acarbose), insulins,meglitinides (e.g., repaglinide), sulfonylureas (e.g., glimepiride,glyburide, and glipizide), thiozolidinediones (e.g. troglitazone,rosiglitazone and pioglitazone), and PPAR-gamma agonists;mineralocorticoid receptor antagonists, such as spironolactone andeplerenone; growth hormone secretagogues; aP2 inhibitors;phosphodiesterase inhibitors, such as PDE III inhibitors (e.g.,cilostazol) and PDE V inhibitors (e.g., sildenafil, tadalafil,vardenafil); protein tyrosine kinase inhibitors; antiinflammatories;antiproliferatives, such as methotrexate, FK506 (tacrolimus, Prograf),mycophenolate mofetil; chemotherapeutic agents; immunosuppressants;anticancer agents and cytotoxic agents (e.g., alkylating agents, such asnitrogen mustards, alkyl sulfonates, nitrosoureas, ethylenimines, andtriazenes); antimetabolites, such as folate antagonists, purineanalogues, and pyrridine analogues; antibiotics, such as anthracyclines,bleomycins, mitomycin, dactinomycin, and plicamycin; enzymes, such asL-asparaginase; farnesyl-protein transferase inhibitors; hormonalagents, such as glucocorticoids (e.g., cortisone),estrogens/antiestrogens, androgens/antiandrogens, progestins, andluteinizing hormone-releasing hormone anatagonists, and octreotideacetate; microtubule-disruptor agents, such as ecteinascidins;microtubule-stabilizing agents, such as pacitaxel, docetaxel, andepothilones A-F; plant-derived products, such as vinca alkaloids,epipodophyllotoxins, and taxanes; and topoisomerase inhibitors;prenyl-protein transferase inhibitors; and cyclosporins; steroids, suchas prednisone and dexamethasone; cytotoxic drugs, such as azathiprineand cyclophosphamide; TNF-alpha inhibitors, such as tenidap; anti-TNFantibodies or soluble TNF receptor, such as etanercept, rapamycin, andleflunimide; and cyclooxygenase-2 (COX-2) inhibitors, such as celecoxiband rofecoxib; and miscellaneous agents such as, hydroxyurea,procarbazine, mitotane, hexamethylmelamine, gold compounds, platinumcoordination complexes, such as cisplatin, satraplatin, and carboplatin.

Thus, in another aspect, certain embodiments provide methods fortreating immune function-mediated disorders in a human or animal subjectin need of such treatment comprising administering to said subject anamount of a compound disclosed herein effective to reduce or preventsaid disorder in the subject, in combination with at least oneadditional agent for the treatment of said disorder that is known in theart. In a related aspect, certain embodiments provide therapeuticcompositions comprising at least one compound disclosed herein incombination with one or more additional agents for the treatment ofimmune function-mediated disorders.

General Synthetic Methods for Preparing Compounds

Isotopic hydrogen can be introduced into a compound as disclosed hereinby synthetic techniques that employ deuterated reagents, wherebyincorporation rates are pre-determined; and/or by exchange techniques,wherein incorporation rates are determined by equilibrium conditions,and may be highly variable depending on the reaction conditions.Synthetic techniques, where tritium or deuterium is directly andspecifically inserted by tritiated or deuterated reagents of knownisotopic content, may yield high tritium or deuterium abundance, but canbe limited by the chemistry required. Exchange techniques, on the otherhand, may yield lower tritium or deuterium incorporation, often with theisotope being distributed over many sites on the molecule.

The compounds as disclosed herein can be prepared by methods known toone of skill in the art and routine modifications thereof, and/orfollowing procedures similar to those described in the Example sectionherein and routine modifications thereof, and/or procedures found inWennerberg et al., Org. Proc. Res. & Dev. 2007, 11(4), 674-80; Wang etal., Bioorganic & Medicinal Chemistry Letters 2007, 17(10), 2817-2822;Jansson et al., J. Org. Chem. 2006, 71(4), 1658-67; Joensson et al., J.Med. Chem. 2004, 47(8), 2075-88; US 2007/088050; US 2005/215586; US2005/192315; US 2004/034227; WO 2005/74899; WO 2003106424; and WO1999/55678, which are hereby incorporated in their entirety, andreferences cited therein and routine modifications thereof. Compounds asdisclosed herein can also be prepared as shown in any of the followingschemes and routine modifications thereof.

The following schemes can be used to practice the present invention. Anyposition shown as hydrogen may be optionally substituted with deuterium.

Compound 1 is reacted with an appropriate chloroformate or phosgeneequivalent, such as isopropyl carbonochloridate, in the presence of anappropriate dehydrating agent, such as acetyl chloride, in anappropriate solvent, such as 1,4-dioxane, at an elevated temperature togive compound 2. Compound 2 is reacted with compound 3 in the presenceof an appropriate base, such as sodium hydride, in an appropriatesolvent, such as dimethylformamide, under an inert atmosphere, such asnitrogen, to give compound 4. Compound 4 is reacted with an appropriatemalonate derivative, such as diethyl malonate, in the presence of anappropriate base, such as sodium hydride, in an appropriate solvent,such as dimethylformamide, at an elevated temperature, to give compound5. Compound 5 is reacted with compound 6 in an appropriate solvent, suchas n-heptane, at an elevated temperature, to give compound 7 of FormulaI.

Deuterium can be incorporated to different positions synthetically,according to the synthetic procedures as shown in Scheme I, by usingappropriate deuterated intermediates. For example, to introducedeuterium at one or more positions of R₄-R₆, compound 1 with thecorresponding deuterium substitutions can be used. To introducedeuterium at one or more positions of R₁-R₃, compound 3 with thecorresponding deuterium substitutions can be used. To introducedeuterium at one or more positions of R₈-R₁₇, compound 6 with thecorresponding deuterium substitutions can be used.

Deuterium can be incorporated to various positions having anexchangeable proton, such as the hydroxyl O—H, via proton-deuteriumequilibrium exchange. For example, to introduce deuterium at R₇, thisproton may be replaced with deuterium selectively or non-selectivelythrough a proton-deuterium exchange method known in the art.

The invention is further illustrated by the following examples. AllIUPAC names were generated using CambridgeSoft's ChemDraw 10.0.

EXAMPLE 1 Sodium5-chloro-3-(ethyl(phenyl)carbamoyl)-1-methyl-2-oxo-1,2-dihydroquinolin-4-olate

Step 1

5-Chloro-1H-benzo[d][1,3]oxazine-2,4-dione: Under an atmosphere ofnitrogen, isopropyl carbonochloridate (50 ml, 4.50 equiv) was addeddropwise to a suspension of 2-amino-6-chlorobenzoic acid (20 g, 116.56mmol, 1.00 equiv) in 1,4-dioxane (150 ml). The resulting solution wasmaintained at about 90° C. for 30 minutes, and then cooled to about 50°C. Acetyl chloride (50 ml, 6.00 equiv) was added in one portion, and thesolution was maintained at about 50° C. for about 30 minutes. Theresulting solids were collected by filtration and purified by silica gelchromatography (ethyl acetate/petroleum ether 10:1) to afford the titleproduct as a gray white solid (17.6 g, yield: 76%).

Step 2

5-Chloro-1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione: Under an atmosphereof nitrogen, 5-chloro-1H-benzo[d][1,3]oxazine-2,4-dione (10 g, 50.61mmol, 1.00 equiv) was dissolved in N,N-dimethylformamide (100 ml) atabout 5° C. Sodium hydride (2.8 g, 121.5 mmol, 2.4 equiv) and methyliodide (5.7 ml, 2 equiv) were then added, and the resulting mixture wasstirred at ambient temperature for about 16 hours. The mixture waspurged with nitrogen for about 1 hour to give the title product as ayellow solid, which was used directly in the next step without anypurification.

Step 3

Ethyl5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate:Sodium hydride (1.9 g, 79.17 mmol, 1.60 equiv) was added in severalportions to the mixture of5-chloro-1-methyl-1H-benzo[d][1,3]oxazine-2,4-dione inN,N-dimethylformamide from Step 2. Diethyl malonate (7.7 g, 48.07 mmol,1.00 equiv) was then added dropwise to the stirred mixture over a periodof about 30 minutes. The resulting solution was stirred at about 85° C.for about 1 hour, water (800 ml) was added, and the pH of the solutionwas adjusted to 2 with a solution of hydrochloric acid (5 mol/L). Theresulting crude product was collected by filtration and thenre-crystallized in ethanol to give the title product as a light yellowsolid (2.5 g, yield: 18% 2 steps).

Step 4

5-Chloro-N-ethyl-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide:N-ethylbenzenamine (430 mg, 3.55 mmol, 2.00 equiv) was added dropwise toethyl5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate(500 mg, 1.78 mmol, 1.00 equiv) dissolved in heptane (10 ml). Theresulting mixture was heated about 100° C. and the volatiles wereremoved by distillation over a period of about 7 hours. After cooling toambient temperature, the resulting crystals were collected byfiltration, washed with heptane, and purified by silica gelchromatography (ethyl acetate/petroleum ether 1:3) to afford the titleproduct as a white solid (0.38 g, yield: 60%).

Step 5

Sodium5-chloro-3-(ethyl(phenyl)carbamoyl)-1-methyl-2-oxo-1,2-dihydroquinolin-4-olate:The pH value of a solution of5-chloro-N-ethyl-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide(170 mg, 0.48 mmol, 1.00 equiv) in ethanol (5 ml) was adjusted to 9-10with a solution of 5M sodium hydroxide. The mixture then was stirred forabout 30 min at ambient temperature. The resulting solids were collectedby filtration and washed with ethanol to give the title compound as awhite solid (70 mg, yield: 39%). ¹H NMR (300 MHz, DMSO) δ: 6.84˜7.31 (m,8H), 3.68 (q, 2H), 3.34 (s, 3H), 1.02 (t, 3H). LC-MS: m/z=357 (M-Na⁺2H)⁺

EXAMPLE 2 Sodium5-chloro-3-(ethyl(phenyl)carbamoyl)-1-d₃-methyl-2-oxo-1,2-dihydroquinolin-4-olate

Step 1

d₃-Ethyl5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate:The procedure of Example 1, Step 2 was followed, but substitutingd₃-methyl iodide for methyl iodide. The resulting product, a yellowsolid, was used directly in the next step without any purification.

Step 3

d₃-Ethyl5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate:The procedure of Example 1, Step 3 was followed, but substitutingd₃-ethyl5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate forethyl5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate.The title product was isolated as a yellow solid (5.8 g, yield: 57% 2steps).

Step 4

d₃-5-Chloro-N-ethyl-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide:The procedure of Example 1, Step 4 was followed, but substitutingd₃-ethyl5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate forethyl5-chloro-4-hydroxy-1-methyl-2-oxo-1,2-dihydroquinoline-3-carboxylate.The title product was isolated as a white solid (1.0 g, yield: 79%).

Step 5

Sodium5-chloro-3-(ethyl(phenyl)carbamoyl)-1-d₃-methyl-2-oxo-1,2-dihydroquinolin-4-olate:The procedure of Example 1, Step 5 was followed, but substitutingd₃-5-chloro-N-ethyl-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamidefor5-chloro-N-ethyl-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide.The title product was isolated as a white solid (0.17 g, yield: 80%). ¹HNMR (300 MHz, DMSO) δ: 6.83˜7.32 (m, 8H), 3.68 (q, 2H), 1.03 (t, 3H).LC-MS: m/z=360 (M-Na⁺2H)⁺.

EXAMPLE 3 Sodium5-chloro-3-(d₅-ethyl(phenyl)carbamoyl)-1-methyl-2-oxo-1,2-dihydroquinolin-4-olate

Step 1

tert-butyl phenylcarbamate: Aniline (2.3 g, 25 mmol, 1 equiv) wasdissolved in tetrahydrofuran (25 ml) at about 5° C. A solution ofdi-tert-butyl dicarbonate (6.0 g, 27.5 mmol) in tetrahydrofuran (10 ml)was added to the solution, and the resulting mixture was heated atreflux for about 2 hours. The solvent was removed in vacuo and theresulting residue was dissolved in ethyl acetate (50 ml). The resultingsolution was washed with a 1M citric acid solution (2×50 ml) and brine(1×50 ml). The organic phase was dried over sodium sulfate andevaporated in vacuo to give the title product as a white solid (4.3 g,yield: 83%).

Step 2

d₅-Ethyl-phenyl-carbamic acid tert-butyl ester: Potassium2-methylpropan-2-olate (790 mg, 7.05 mmol, 2.50 equiv) and d₅-iodoethane(500 mg, 3.11 mmol, 1.10 equiv) were added to tert-butyl phenylcarbamate(540 mg, 2.80 mmol, 1.00 equiv) dissolved in N,N-dimethylformamide (100mL). The resulting mixture was stirred at about 55° C. for about 16hours, and then deuterium oxide was added (10 ml). The pH of the mixturewas then adjusted to about 6-7 with 1N hydrochloric acid. Standardextractive workup with ethyl acetate gave the title product as a cruderesidue, which was used in the next step without further purification.

Step 3

N-d₅-Ethylbenzenamine: Over a period of 1 hour and maintaining thetemperature at around 25° C., hydrochloric gas was introduced tod₅-ethyl-phenyl-carbamic acid tert-butyl ester dissolved in ethylacetate (5 ml). The pH of the solution was then adjusted to 6-7 with asodium hydroxide solution (10 mol/L). Standard extractive workup withethyl acetate gave the title product as yellow oil (0.33 g, yield: 93%).

Step 4

5-Chloro-N-d₅-ethyl-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide:The procedure of Example 1, Step 4 was followed, but substitutingN-d₅-ethylbenzenamine for N-ethylbenzenamine. The title product wasisolated as a white solid (0.4 g, yield: 58%).

Step 5

Sodium5-chloro-3-(d₅-ethyl(phenyl)carbamoyl)-1-methyl-2-oxo-1,2-dihydroquinolin-4-olate:The procedure of Example 1, Step 5 was followed, but substituting5-chloro-N-d₅-ethyl-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamidefor5-chloro-N-ethyl-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide.The title product was isolated as a white solid (90 mg, yield: 40.5%).¹H NMR (300 MHz, DMSO) δ: 6.84˜7.32 (m, 8H), 3.34 (s, 3H) LC-MS: m/z=362(M-Na+2H)⁺.

EXAMPLE 4 Sodium5-chloro-3-(d₅-ethyl(phenyl)carbamoyl)-1-d₃-methyl-2-oxo-1,2-dihydroquinolin-4-olate

Step 1

5-chloro-N-d₈-ethyl-4-hydroxy-1-methyl-2-oxo-N-phenyl-1,2-dihydroquinoline-3-carboxamide:The procedure of Example 2, Step 4 was followed, but substitutingN-d₅-ethylbenzenamine for N-ethylbenzenamine. The title product wasisolated as a white solid.

Step 2

Sodium5-chloro-3-(d₈-ethyl(phenyl)carbamoyl)-1-d₃-methyl-2-oxo-1,2-dihydroquinolin-4-olate:The procedure of Example 2, Step 5 was followed, but substitutingN-d₅-ethylbenzenamine for N-ethylbenzenamine. The title product wasisolated as a white solid (0.1 g, yield: 70%). ¹H NMR (300 MHz, DMSO) δ:6.83˜7.31 (m, 8H). LC-MS: m/z=365 (M-Na⁺2H)⁺.

The following compounds can generally be made using the methodsdescribed above. It is expected that these compounds when made will haveactivity similar to those described in the examples above.

Changes in the metabolic properties of the compounds disclosed herein ascompared to their non-isotopically enriched analogs can be shown usingthe following assays. Compounds listed above which have not yet beenmade and/or tested are predicted to have changed metabolic properties asshown by one or more of these assays as well.

Biological Activity Assays

In Vitro Liver Microsomal Stability Assay

Liver microsomal stability assays are conducted at 2 mg per mL livermicrosome protein with an NADPH-generating system in 2% sodiumbicarbonate (2.2 mM NADPH, 25.6 mM glucose 6-phosphate, 6 units per mLglucose 6-phosphate dehydrogenase and 3.3 mM magnesium chloride). Testcompounds are prepared as solutions in 20% acetonitrile-water and addedto the assay mixture (final assay concentration 5 microgram per mL) andincubated at 37° C. Final concentration of acetonitrile in the assayshould be <1%. Aliquots (50 μL) are taken out at times 0, 30, 60, 90,and 120 minutes, and diluted with ice cold acetonitrile (200 μL) to stopthe reactions. Samples are centrifuged at 12,000 RPM for 10 minutes toprecipitate proteins. Supernatants are transferred to microcentrifugetubes and stored for LC/MS/MS analysis of the degradation half-life ofthe test compounds. It has thus been found that certaindeuterium-enriched compounds disclosed herein that have been tested inthis assay showed an increased degradation half-life as compared to thenon-isotopically enriched drug. In certain embodiments, the increase indegradation half-life is at least 5%; at least 10%; at least 15%; atleast 20%; at least 30%; at least 40%; at least 50%; at least 60%; atleast 70%; at least 80%; at least 90%; or at least 100%.

In Vitro Metabolism Using Human Cytochrome P₄₅₀ Enzymes

The cytochrome P₄₅₀ enzymes are expressed from the corresponding humancDNA using a baculovirus expression system (BD Biosciences, San Jose,Calif.). A 0.25 milliliter reaction mixture containing 0.8 milligramsper milliliter protein, 1.3 millimolar NADP⁺, 3.3 millimolarglucose-6-phosphate, 0.4 U/mL glucose-6-phosphate dehydrogenase, 3.3millimolar magnesium chloride and 0.2 millimolar of a compound ofFormula I, the corresponding non-isotopically enriched compound orstandard or control in 100 millimolar potassium phosphate (pH 7.4) isincubated at 37° C. for 20 min. After incubation, the reaction isstopped by the addition of an appropriate solvent (e.g., acetonitrile,20% trichloroacetic acid, 94% acetonitrile/6% glacial acetic acid, 70%perchloric acid, 94% acetonitrile/6% glacial acetic acid) andcentrifuged (10,000 g) for 3 min. The supernatant is analyzed byHPLC/MS/MS.

Cytochrome P₄₅₀ Standard CYP1A2 Phenacetin CYP2A6 Coumarin CYP2B6[¹³C]-(S)-mephenytoin CYP2C8 Paclitaxel CYP2C9 Diclofenac CYP2C19[¹³C]-(S)-mephenytoin CYP2D6 (+/−)-Bufuralol CYP2E1 Chlorzoxazone CYP3A4Testosterone CYP4A [¹³C]-Lauric acidMonoamine Oxidase A Inhibition and Oxidative Turnover

The procedure is carried out using the methods described by Weyler,Journal of Biological Chemistry 1985, 260, 13199-13207, which is herebyincorporated by reference in its entirety. Monoamine oxidase A activityis measured spectrophotometrically by monitoring the increase inabsorbance at 314 nm on oxidation of kynuramine with formation of4-hydroxyquinoline. The measurements are carried out, at 30° C., in 50mM sodium phosphate buffer, pH 7.2, containing 0.2% Triton X-100(monoamine oxidase assay buffer), plus 1 mM kynuramine, and the desiredamount of enzyme in 1 mL total volume.

Monoamine Oxidase B Inhibition and Oxidative Turnover

The procedure is carried out as described in Uebelhack,Pharmacopsychiatry 1998, 31(5), 187-192, which is hereby incorporated byreference in its entirety.

Determining Laquinimod in Plasma by Coupled-Column Liquid Chromatographywith Ultraviolet Absorbance Detection

The procedure is carried out as described in Edman, et al., Journal ofChromatography, B: Analytical Technologies in the Biomedical and LifeSciences 2003, 785(2), which is hereby incorporated by reference in itsentirety.

Determining Laquinimod in Human Plasma by Liquid Chromatography/TandemMass Spectrometry

The procedure is carried out as described in Sennbro, et al., RapidCommunications in Mass Spectrometry 2006, 20(22), 3313-3318, which ishereby incorporated by reference in its entirety.

Measuring Laquinimod's Effect on Th1/Th2 Balance and Th3 Cytokine andTGF-β Cytokine Production in Lewis Rats.

The procedure is carried out as described in Yang, et al., Journal ofNeuroimmunology 2004, 156(1-2), 3-9, which is hereby incorporated byreference in its entirety.

Experimental Autoimmune Encephalomyelitis Model

The procedure is carried out as described in Karussis et al., Ann.Neurol. 1993, 34, 654-660, which is hereby incorporated by reference inits entirety.

From the foregoing description, one skilled in the art can ascertain theessential characteristics of this invention, and without departing fromthe spirit and scope thereof, can make various changes and modificationsof the invention to adapt it to various usages and conditions.

1. A compound of structural Formula I

or a salt thereof, wherein: R₁-R₁₇ are independently selected from thegroup consisting of hydrogen and deuterium; and at least one of R1-R17is deuterium.
 2. The compound as recited in claim 1 wherein at least oneof R₁-R₁₇ independently has deuterium enrichment of no less than about10%.
 3. The compound as recited in claim 1 wherein at least one ofR₁-R₁₇ independently has deuterium enrichment of no less than about 50%.4. The compound as recited in claim 1 wherein at least one of R₁-R₁₇independently has deuterium enrichment of no less than about 90%.
 5. Thecompound as recited in claim 1 wherein at least one of R₁-R₁₇independently has deuterium enrichment of no less than about 98%.
 6. Thecompound as recited in claim 1 wherein said compound has a structuralformula selected from the group consisting of:


7. The compound as recited in claim 1 wherein said compound has astructural formula selected from the group consisting of


8. The compound as recited in claim 7 wherein each position representedas D has deuterium enrichment of no less than about 10%.
 9. The compoundas recited in claim 7 wherein each position represented as D hasdeuterium enrichment of no less than about 50%.
 10. The compound asrecited in claim 7 wherein each position represented as D has deuteriumenrichment of no less than about 90%.
 11. The compound as recited inclaim 7 wherein each position represented as D has deuterium enrichmentof no less than about 98%.
 12. The compound as recited in claim 7wherein said compound has the structural formula:


13. The compound as recited in claim 7 wherein said compound has thestructural formula:


14. The compound as recited in claim 7 wherein said compound has thestructural formula:


15. A pharmaceutical composition comprising a compound as recited inclaim 1 together with a pharmaceutically acceptable carrier.
 16. Amethod of treatment of a immune function-mediated disorder comprisingthe administration of a therapeutically effective amount of a compoundas recited in claim 1 to a patient in need thereof.
 17. The method asrecited in claim 16 wherein said disorder is multiple sclerosis andautoimmune disorders.
 18. The method as recited in claim 16 furthercomprising the administration of an additional therapeutic agent. 19.The method as recited in claim 18 wherein said additional therapeuticagent is selected from the group consisting of immunomodulators andcyclosporins.
 20. The method as recited in claim 19 wherein saidimmunomodulator is selected from the group consisting of filgrastim,molgramostim, sargramostim, lenograstim, ancestim, pegfilgrastim,interferon gamma, interferon alpha-2a, interferon alpha-2b, interferonalpha-n1, interferon beta-1a, interferon beta-1b, interferon alphacon-1,peginterferon alpha-2b, peginterferon alpha-2a, interferon omega,aldesleukin, oprelvekin, lentinan, roquinimex, BCG vaccine, pegademase,pidotimod, Poly I:C, Poly ICLC, thymopentin, immunocyanin, tasonermin,melanoma vaccine, glatiramer acetate, histamine dihydrochloride,mifamurtide, plerixefor, muromonab-CD3, antilymphocyte immunoglobulin(horse), antithymocyte immunoglobulin (rabbit), mycophenolic acid,sirolimus, leflunomide, alefacept, everolimus, gusperimus, efalizumab,abetimus, natalizumab, abatacept, eculizumab, etanercept, infliximab,afelimomab, adalimumab, certolizumab pegol, daclizumab, basiliximab,anakinra, ciclosporin, tacrolimus, azathioprine, thalidomide,methotrexate, and lenalidomide.
 21. The method as recited in claim 16,further resulting in least one effect selected from the group consistingof: a. decreased inter-individual variation in plasma levels of saidcompound or a metabolite thereof as compared to the non-isotopicallyenriched compound; b. increased average plasma levels of said compoundper dosage unit thereof as compared to the non-isotopically enrichedcompound; c. decreased average plasma levels of at least one metaboliteof said compound per dosage unit thereof as compared to thenon-isotopically enriched compound; d. increased average plasma levelsof at least one metabolite of said compound per dosage unit thereof ascompared to the non-isotopically enriched compound; and e. an improvedclinical effect during the treatment in said subject per dosage unitthereof as compared to the non-isotopically enriched compound.
 22. Themethod as recited in claim 16, further resulting in at least two effectsselected from the group consisting of: a. decreased inter-individualvariation in plasma levels of said compound or a metabolite thereof ascompared to the non-isotopically enriched compound; b. increased averageplasma levels of said compound per dosage unit thereof as compared tothe non-isotopically enriched compound; c. decreased average plasmalevels of at least one metabolite of said compound per dosage unitthereof as compared to the non-isotopically enriched compound; d.increased average plasma levels of at least one metabolite of saidcompound per dosage unit thereof as compared to the non-isotopicallyenriched compound; and e. an improved clinical effect during thetreatment in said subject per dosage unit thereof as compared to thenon-isotopically enriched compound.
 23. The method as recited in claim16, wherein the method effects a decreased metabolism of the compoundper dosage unit thereof by at least one polymorphically-expressedcytochrome P450 isoform in the subject, as compared to the correspondingnon-isotopically enriched compound.
 24. The method as recited in claim23, wherein the cytochrome P450 isoform is selected from the groupconsisting of CYP2C8, CYP2C9, CYP2C19, and CYP2D6.
 25. The method asrecited claim 16, wherein said compound is characterized by decreasedinhibition of at least one cytochrome P 450 or monoamine oxidase isoformin said subject per dosage unit thereof as compared to thenon-isotopically enriched compound.
 26. The method as recited in claim25, wherein said cytochrome P450 or monoamine oxidase isoform isselected from the group consisting of CYPIAI, CYPIA2, CYPIBI, CYP2A6,CYP2A13, CYP2B6, CYP2C8, CYP2C9, CYP2CI8, CYP2CI9, CYP2D6, CYP2EI,CYP2GI, CYP2J2, CYP2RI, CYP2SI, CYP3A4, CYP3A5, CYP3A5PI, CYP3A5P2,CYP3A7, CYP4AII, CYP4BI, CYP4F2, CYP4F3, CYP4F8, CYP4Fll, CYP4FI2,CYP4XI, CYP4ZI, CYP5AI, CYP7AI, CYP7BI, CYP8AI, CYP8BI, CYPIIAI,CYPIIBI, CYPIIB2, CYPI7, CYPI9, CYP21, CYP24, CYP26AI, CYP26BI, CYP27AI,CYP27BI, CYP39, CYP46, CYP51, MAOA, and MAOB.
 27. The method as recitedin claim 16, wherein the method reduces a deleterious change in adiagnostic hepatobiliary function endpoint, as compared to thecorresponding non-isotopically enriched compound.
 28. The method asrecited in claim 27, wherein the diagnostic hepatobiliary functionendpoint is selected from the group consisting of alanineaminotransferase (“ALT”), serum glutamic-pyruvic transaminase (“SGPT”),aspartate aminotransferase (“AST,” “SGOT”), ALT/AST ratios, serumaldolase, alkaline phosphatase (“ALP”), ammonia levels, bilirubin,gamma-glutamyl transpeptidase (“GGTP,” “γ-GTP,” “GGT”), leucineaminopeptidase (“LAP”), liver biopsy, liver ultrasonography, livernuclear scan, 5′-nucleotidase, and blood protein.